Hepatitis E virus (HEV) is the major etiologic agent for enterically transmitted non-A, non-B hepatitis. Hepatitis E represents more than half of the acute viral hepatitis cases occurring in developing countries where large epidemics affect young to middle-aged adults. Systematic studies on HEV replication and pathogenesis are needed to guide diagnosis and prevention. No current method is available to grow HEV in culture. The overall goal of this proposal is to identify an in vitro infectious HEV molecule and to establish a cell culture system amenable to supporting viral replication. A full-length cDNA clone and in vitro-synthesized RNA transcripts of HEV will be tested for their respective abilities to infect susceptible cells in culture. To confirm in vitro infection, we will examine RNA- and DNA-transfected cells for the presence of replicative RNA species as well as authentic viral proteins. With the establishment of a cell culture system for HEV infection, we will, in future phase studies, investigate its potential utilities to provide useful reagents for the development of diagnostics and vaccine. It will also provide a model to study HEV cellular pathogenesis and thus potential ways to interfere with the process.